Cefoperazone-treated Mouse Model of Clinically-relevant <em>Clostridium difficile</em> Strain R20291

Clostridium difficile is an anaerobic, gram-positive, spore-forming enteric pathogen that is associated with increasing morbidity and mortality and consequently poses an urgent threat to public health. Recurrence of a C. difficile infection (CDI) after successful treatment with antibiotics is high, occurring in 20–30% of patients, thus necessitating the discovery of novel therapeutics against this pathogen. Current animal models of CDI result in high mortality rates and thus do not approximate the chronic, insidious disease manifestations seen in humans with CDI. To evaluate therapeutics against C. difficile, a mouse model approximating human disease utilizing a clinically-relevant strain is needed. This protocol outlines the cefoperazone mouse model of CDI using a clinically-relevant and genetically-tractable strain, {"type":"entrez-nucleotide","attrs":{"text":"R20291","term_id":"774925","term_text":"R20291"}}R20291. Techniques for clinical disease monitoring, C. difficile bacterial enumeration, toxin cytotoxicity, and histopathological changes throughout CDI in a mouse model are detailed in the protocol. Compared to other mouse models of CDI, this model is not uniformly lethal at the dose administered, allowing for the observation of a prolonged clinical course of infection concordant with the human disease. Therefore, this cefoperazone mouse model of CDI proves a valuable experimental platform to assess the effects of novel therapeutics on the amelioration of clinical disease and on the restoration of colonization resistance against C. difficile

Mutations in the E2 glycoprotein and the 3\u27 untranslated region enhance chikungunya virus virulence in mice

Chikungunya virus (CHIKV) is a mosquito-transmitted alphavirus that causes debilitating musculoskeletal pain and inflammation and can persist for months to years after acute infection. Although studies of humans and experimentally infected animals suggest that CHIKV infection persists in musculoskeletal tissues, the mechanisms for this remain poorly understood. To evaluate this further, we isolated CHIKV from the serum of persistently infected Rag1 -/- mice at day 28. When inoculated into naive wild-type (WT) mice, this persistently circulating CHIKV strain displayed a capacity for earlier dissemination and greater pathogenicity than the parental virus. Sequence analysis revealed a nonsynonymous mutation in the E2 glycoprotein (E2 K200R) and a deletion within the 3' untranslated region (3'-UTR). The introduction of these changes into the parental virus conferred enhanced virulence in mice, although primary tropism for musculoskeletal tissues was maintained. The E2 K200R mutation was largely responsible for enhanced viral dissemination and pathogenicity, although these effects were augmented by the 3'- UTR deletion. Finally, studies with Irf3/Irf7 -/- and Ifnar1 -/- mice suggest that the E2 K200R mutation enhances viral dissemination from the site of inoculation independently of interferon regulatory factor 3 (IRF3)-, IRF7-, and IFNAR1-mediated responses. As our findings reveal viral determinants of CHIKV dissemination and pathogenicity, their further study should help to elucidate host-virus interactions that determine acute and chronic CHIKV infection

Molecular and Functional Mapping of EED Motifs Required for PRC2-Dependent Histone Methylation

Polycomb Group (PcG) proteins represent a conserved family of developmental regulators that mediate heritable transcriptional silencing by modifying chromatin states. One PcG complex, the PRC2 complex, is composed of several proteins, including the histone H3 lysine 27 (H3K27) methyltransferase EZH2 and the WD-repeat protein EED. Histone H3K27 can be mono- (H3K27me1), di- (H3K27me2), or trimethylated (H3K27me3). However, it remains unclear what regulates the number of methyl groups added to H3K27 in a particular nucleosome. In mammalian cells, EED is present as four distinct isoforms, which are believed to be produced by utilizing four distinct, in-frame translation start sites in a common Eed mRNA. A mutation that disables all four EED isoforms produces defects in H3K27 methylation.1 To assess the roles of individual EED isoforms in H3K27 methylation, we first characterized three of the four EED isoform start sites and then demonstrated that individual isoforms are not necessary for H3K27me1, H3K27me2, or H3K27me3. Instead, we show that the core WD-40 motifs and the histone binding region of EED alone are sufficient for the generation of all three marks, demonstrating that EED isoforms do not control the number of methyl groups added to H3K27

To tweet or not to tweet about schizophrenia systematic reviews (TweetSz): study protocol for a randomised controlled trial

Introduction: The Cochrane Schizophrenia Group (CSzG) has produced and maintained systematic reviews of effects of interventions for schizophrenia and related illness. Each review has a Plain Language Summary (PLS), for those without specialised knowledge, and an abstract, which are freely available from The Cochrane Library (https://summaries.cochrane.org). Increasingly, evidence is being distributed using social media such as Twitter and Weibo (in China) alongside traditional publications. Methods and analysis: In a prospective two-arm, parallel, open randomised controlled trial with a 1:1 allocation ratio, we will allocate 170 published systematic reviews into the intervention group (tweeting arm/Weibo arm) versus the control group (non-tweeting arm). Reviews will be stratified by baseline access activity, defined as high (≥19 views per week, n=14), medium (4.3 to 18.99 views per week, n=72) or low (<4.3 views per week, n=84), based on Google Analytics, which will also be used for evaluating outcomes. The intervention group will have three tweets daily using Hootsuite with a slightly different accompanying text (written by CEA and AB) and a shortened Uniform Resource Locator (URL) to the PLS: a) The review title as it appears in summaries.cochrane.org, b) A pertinent extract from results or discussion sections of the abstract and c) An intriguing question or pithy statement related to the evidence in the abstract. The primary outcome will be: total number of visits to a PLS in 7 days following the tweet. Secondary outcomes will include % new visits, bounce rate, pages per visit, visit duration, page views, unique page views, time on page, entrances, exiting behaviour and country distribution. Ethics and dissemination: This study does not involve living participants, and uses information available in the public domain. Participants are published systematic reviews, hence, no ethical approval is required. Dissemination will be via Twitter, Weibo and traditional academic means

Toward Personalized Gene Therapy: Characterizing the Host Genetic Control of Lentiviral-Vector-Mediated Hepatic Gene Delivery

The success of lentiviral vectors in curing fatal genetic and acquired diseases has opened a new era in human gene therapy. However, variability in the efficacy and safety of this therapeutic approach has been reported in human patients. Consequently, lentiviral-vector-based gene therapy is limited to incurable human diseases, with little understanding of the underlying causes of adverse effects and poor efficacy. To assess the role that host genetic variation has on efficacy of gene therapy, we characterized lentiviral-vector gene therapy within a set of 12 collaborative cross mouse strains. Lentiviral vectors carrying the firefly luciferase cDNA under the control of a liver-specific promoter were administered to female mice, with total-body and hepatic luciferase expression periodically monitored through 41 weeks post-vector administration. Vector copy number per diploid genome in mouse liver and spleen was determined at the end of this study. We identified major strain-specific contributions to overall success of transduction, vector biodistribution, maximum luciferase expression, and the kinetics of luciferase expression throughout the study. Our results highlight the importance of genetic variation on gene-therapeutic efficacy; provide new models with which to more rigorously assess gene therapy approaches; and suggest that redesigning preclinical studies of gene-therapy methodologies might be appropriate

Knockout of Epstein-Barr Virus BPLF1 Retards B-Cell Transformation and Lymphoma Formation in Humanized Mice

ABSTRACT BPLF1 of Epstein-Barr virus (EBV) is classified as a late lytic cycle protein but is also found in the viral tegument, suggesting its potential involvement at both initial and late stages of viral infection. BPLF1 possesses both deubiquitinating and deneddylating activity located in its N-terminal domain and is involved in processes that affect viral infectivity, viral DNA replication, DNA repair, and immune evasion. A recently constructed EBV BPLF1-knockout (KO) virus was used in conjunction with a humanized mouse model that can be infected with EBV, enabling the first characterization of BPLF1 function in vivo . Results demonstrate that the BPLF1-knockout virus is approximately 90% less infectious than wild-type (WT) virus. Transformation of human B cells, a hallmark of EBV infection, was delayed and reduced with BPLF1-knockout virus. Humanized mice infected with EBV BPLF1-knockout virus showed less weight loss and survived longer than mice infected with equivalent infectious units of WT virus. Additionally, splenic tumors formed in 100% of mice infected with WT EBV but in only 25% of mice infected with BPLF1-KO virus. Morphological features of spleens containing tumors were similar to those in EBV-induced posttransplant lymphoproliferative disease (PTLD) and were almost identical to cases seen in human diffuse large B-cell lymphoma. The presence of EBV genomes was detected in all mice that developed tumors. The results implicate BPLF1 in human B-cell transformation and tumor formation in humanized mice. IMPORTANCE Epstein-Barr virus infects approximately 90% of the world’s population and is the causative agent of infectious mononucleosis. EBV also causes aggressive lymphomas in individuals with acquired and innate immune disorders and is strongly associated with diffuse large B-cell lymphomas, classical Hodgkin lymphoma, Burkitt lymphoma, and nasopharyngeal carcinoma (NPC). Typically, EBV initially infects epithelial cells in the oropharynx, followed by a lifelong persistent latent infection in B-cells, which may develop into lymphomas in immunocompromised individuals. This work is the first of its kind in evaluating the effects of EBV’s BPLF1 in terms of pathogenesis and lymphomagenesis in humanized mice and implicates BPLF1 in B-cell transformation and tumor development. Currently, there is no efficacious treatment for EBV, and therapeutic targeting of BPLF1 may lead to a new path to treatment for immunocompromised individuals or transplant recipients infected with EBV

Evaluation Findings: School and Workplace Strategies 2005-2007

This report is an evaluation developed to provide updates on the progress of the Missouri Foundation for Health’s Tobacco Prevention and Cessation Initiative (TPCI). The report provides an overview of the activities and outcomes regarding school and workplace strategies that occurred between 2005 and 2007.https://openscholarship.wustl.edu/cphss/1099/thumbnail.jp

CD8 + T Cells Control Ross River Virus Infection in Musculoskeletal Tissues of Infected Mice

Ross River virus (RRV), chikungunya virus (CHIKV), and related alphaviruses cause debilitating polyarthralgia and myalgia. Mouse models of RRV and CHIKV have demonstrated a role for the adaptive immune response in the control of these infections. However, questions remain regarding the role for T cells in viral control, including the magnitude, location, and dynamics of CD8+ T cell responses. To address these questions, we generated a recombinant RRV expressing the H-2b-restricted gp33 determinant derived from the glycoprotein (gp) of lymphocytic choriomeningitis virus (LCMV) (“RRV-LCMV”). Utilizing tetramers, we tracked gp33-specific CD8+ T cells during RRV-LCMV infection. We found that acute RRV infection induces activation of CD8+ T cell responses in lymphoid and musculoskeletal tissues that peak from 10 to 14 days post-inoculation (dpi), suggesting that CD8+ T cells contribute to control of acute RRV infection. Mice genetically deficient for CD8+ T cells or wild-type mice depleted of CD8+ T cells had elevated RRV loads in skeletal muscle tissue, but not joint-associated tissues, at 14 dpi, suggesting that the ability of CD8+ T cells to control RRV infection is tissue-dependent. Finally, adoptively transferred T cells were capable of reducing RRV loads in skeletal muscle tissue of Rag1−/− mice, indicating that T cells can contribute to the control of RRV infection in the absence of B cells and antibody. Collectively, these data demonstrate a role for T cells in the control of RRV infection and suggest that the antiviral capacity of T cells is controlled in a tissue-specific manner

An Integrated Framework to Assess Greenwashing

In this paper we examine definitions of ‘greenwashing’ and its different forms, developing a tool for assessing diverse ‘green’ claims made by various actors. Research shows that significant deception and misleading claims exist both in the regulated commercial sphere, as well as in the unregulated non-commercial sphere (e.g., governments, NGO partnerships, international pledges, etc.). Recently, serious concerns have been raised over rampant greenwashing, in particular with regard to rapidly emerging net zero commitments. The proposed framework we developed is the first actionable tool for analysing the quality and truthfulness of such claims. The framework has widespread and unique potential for highlighting efforts that seek to delay or distract real solutions that are urgently needed today to tackle multiple climate and environmental crises. In addition, we note how the framework may also assist in the development of practices and communication strategies that ultimately avoid greenwashing

An Integrated Framework to Assess Greenwashing

Funding: This research was funded by the Department of Political Science at University of Vienna, Austria and the Institute at Brown for Environment and Society, USA, in association with Climate Social Science Network.Peer reviewedPublisher PD